03/24/2022
Optimizing Plasmid Yields in Shake Culture
Blandine Vanbellinghen, Silvia Tejerina, Eppendorf Application Technologies S.A.
Ines Hartmann, Eppendorf SE
Recombinant plasmid DNA is produced in bacterial cultures, mostly in E. coli. Plasmid yield and quality depend on multiple factors, including the insert, the selection of host strain, the vector design, and the methodology chosen for cultivation and downstream purification. Here, we focus on how to optimize E. coli shake flask cultures examining the influence of culture media, vessel design, fill volume, and shaking speed on bacteria and plasmid yields. We illustrate how larger production range can be obtained by using the appropriate combination between a large incubated shaker, optimized culture conditions, and specificallydesigned culture flasks.
Introduction
Plasmids serve as vehicles in genetic engineering either to clone and amplify DNA fragments, such as genes, or express recombinant proteins. Plasmid DNA (pDNA) can be easily genetically manipulated, produced in E. coli in large amounts and a variety of ready-to-use solutions allows easy subsequent downstream purification. Depending on the application, pDNA production ranks from research laboratory scale (up to a few mg) to industrial scales (mg to g scales). Here, we will examine the impact of cultivation with focus on high yield pDNA production in 2.5 L Ultra YieldR flasks.